Intracellular bacteria rely on secretion systems to modulate the host environment and ensure survival. For Coxiella burnetti, the Type IV secretion system (T4SS) translocates bacterial effector proteins into the host cell, creating a hospitable environment critical for productive infection of host cells. While the T4SS is required for intracellular growth, secretion does not begin until Coxiella have been trafficked to lysosome-like compartments - suggesting that restriction of secretion benefits the bacterium and that environmental signals provide the impetus for expression of T4SS and its effectors. Coxiella possess only a few two-component systems, including one (QseBC) related to other systems that activate pathogen secretion in response to the host environment. Indeed, a strain with a transposon insertion disrupting the QseBC operon is neither able to secrete a T4BSS effector nor survive within host cells. The specific aims of this proposed research use genetic and cell biological tools to investigate whether the Coxiella QseBC system senses and responds to the maturing phagosome to initiate expression of the T4SS and its effectors. Aim 1 will use a transcriptional profiling and reporter strains to determine the maximal expression of QseB-regulated genes during both axenic and intracellular growth, correlating expression with vacuolar maturation and medium composition to reveal how and when the QseBC system is activated. Aim 2 will use expression of modified QseB proteins to dysregulate T4SS activity and measure inflammatory reaction to secretion in macrophage-like host cells. Together, Aims 1 and 2 will reveal whether temporal and spatial control of secretion enhances Coxiella's ability to evade host defenses, informing our understanding of acute and chronic Q fever infections. Aim 3 will use the information and experimental tools developed during Aims 1 and 2 to intelligently compare mRNA profiles of strains with or without functional QseB. Aim 3 will reveal the full suite of genes regulated by QseB and their relative expression, as well as provide the first whole-transcriptome analysis of Coxiella burnetii. Together, these complementary aims will provide tools to enhance research across the Coxiella field, greatly expand our understanding of Coxiella gene regulation, and suggest fertile avenues for future research into the biology and genetics of an understudied human and animal pathogen.